Summary Funds are requested to purchase a direct electron detector and advanced operating software for a Titan Krios transmission electron microscope. The new generation of direct electron detectors replace scintillators and CCDs for recording EM images, and offer a highly dramatic improvement in both sensitivity and resolution. The Falcon II camera requested will also offer a three-fold increase in the data collection rate over our existing CCD camera, but the main advantage comes from the huge increase in the signal-to-noise ratio and the greatly improved point spread function. While the three-fold increase in the rate of acquiring images could be met with our existing CCD camera by simply acquiring images over a three-fold longer period, no amount of images acquired with the CCD would ever recover the high resolution information that is either completely absent in the CCD images or buried in the noise. These advances with the direct electron detector will allow us to image a large number of specimens at an unprecedented resolution, greatly aiding a number of NIH-supported projects. Four Major Users (Egelman, Yeager, Stewart and Zhang) and a number of Minor Users will use the Titan Krios as a shared facility. The incredible throughput of this microscope with the proposed camera (~ 2,000 images per day), combined with the robotic operation of the microscope on a 24/7 schedule, means that data analysis will become the rate-limiting step and not image acquisition. This high resolution, high throughput mode of imaging will largely transform EM. Thus, our Krios can truly function as a facility much like synchrotrons do in providing a unique service to the structural biology community. The specimens that will be examined at high resolution on the Krios with the proposed detector range from viruses and bacterial pili to complexes involved in the innate immune recognition of foreign RNA, allowing for conceptual advances in understanding many systems that are important to human health.